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Isolation and characterisation of human pulmonary microvascular endothelial cells from patients with severe emphysema
BACKGROUND: Loss of the pulmonary microvasculature in the pathogenesis of emphysema has been put forward as a credible alternative to the classical inflammatory cell driven proteolysis hypothesis. Mechanistic studies in this area have to date employed animal models, immortalised cell lines, primary...
Autores principales: | , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
BioMed Central
2013
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3599007/ https://www.ncbi.nlm.nih.gov/pubmed/23425195 http://dx.doi.org/10.1186/1465-9921-14-23 |
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author | Mackay, Laura S Dodd, Sara Dougall, Iain G Tomlinson, Wendy Lordan, James Fisher, Andrew J Corris, Paul A |
author_facet | Mackay, Laura S Dodd, Sara Dougall, Iain G Tomlinson, Wendy Lordan, James Fisher, Andrew J Corris, Paul A |
author_sort | Mackay, Laura S |
collection | PubMed |
description | BACKGROUND: Loss of the pulmonary microvasculature in the pathogenesis of emphysema has been put forward as a credible alternative to the classical inflammatory cell driven proteolysis hypothesis. Mechanistic studies in this area have to date employed animal models, immortalised cell lines, primary endothelial cells isolated from large pulmonary arteries and non-pulmonary tissues and normal human pulmonary microvascular endothelial cells. Although these studies have increased our understanding of endothelial cell function, their relevance to mechanisms in emphysema is questionable. Here we report a successful technique to isolate and characterise primary cultures of pulmonary microvascular endothelial cells from individuals with severe emphysema. METHODS: A lobe of emphysematous lung tissue removed at the time of lung transplantation surgery was obtained from 14 patients with severe end-stage disease. The pleura, large airways and large blood vessels were excised and contaminating macrophages and neutrophils flushed from the peripheral lung tissue before digestion with collagenase. Endothelial cells were purified from the cell mixture via selection with CD31 and UEA-1 magnetic beads and characterised by confocal microscopy and flow cytometry. RESULTS: Successful isolation was achieved from 10 (71%) of 14 emphysematous lungs. Endothelial cells exhibited a classical cobblestone morphology with high expression of endothelial cell markers (CD31) and low expression of mesenchymal markers (CD90, αSMA and fibronectin). E-selectin (CD62E) was inducible in a proportion of the endothelial cells following stimulation with TNFα, confirming that these cells were of microvascular origin. CONCLUSIONS: Emphysematous lungs removed at the time of transplantation can yield large numbers of pulmonary microvasculature endothelial cells of high purity. These cells provide a valuable research tool to investigate cellular mechanisms in the pulmonary microvasculature relevant to the pathogenesis of emphysema. |
format | Online Article Text |
id | pubmed-3599007 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2013 |
publisher | BioMed Central |
record_format | MEDLINE/PubMed |
spelling | pubmed-35990072013-03-17 Isolation and characterisation of human pulmonary microvascular endothelial cells from patients with severe emphysema Mackay, Laura S Dodd, Sara Dougall, Iain G Tomlinson, Wendy Lordan, James Fisher, Andrew J Corris, Paul A Respir Res Research BACKGROUND: Loss of the pulmonary microvasculature in the pathogenesis of emphysema has been put forward as a credible alternative to the classical inflammatory cell driven proteolysis hypothesis. Mechanistic studies in this area have to date employed animal models, immortalised cell lines, primary endothelial cells isolated from large pulmonary arteries and non-pulmonary tissues and normal human pulmonary microvascular endothelial cells. Although these studies have increased our understanding of endothelial cell function, their relevance to mechanisms in emphysema is questionable. Here we report a successful technique to isolate and characterise primary cultures of pulmonary microvascular endothelial cells from individuals with severe emphysema. METHODS: A lobe of emphysematous lung tissue removed at the time of lung transplantation surgery was obtained from 14 patients with severe end-stage disease. The pleura, large airways and large blood vessels were excised and contaminating macrophages and neutrophils flushed from the peripheral lung tissue before digestion with collagenase. Endothelial cells were purified from the cell mixture via selection with CD31 and UEA-1 magnetic beads and characterised by confocal microscopy and flow cytometry. RESULTS: Successful isolation was achieved from 10 (71%) of 14 emphysematous lungs. Endothelial cells exhibited a classical cobblestone morphology with high expression of endothelial cell markers (CD31) and low expression of mesenchymal markers (CD90, αSMA and fibronectin). E-selectin (CD62E) was inducible in a proportion of the endothelial cells following stimulation with TNFα, confirming that these cells were of microvascular origin. CONCLUSIONS: Emphysematous lungs removed at the time of transplantation can yield large numbers of pulmonary microvasculature endothelial cells of high purity. These cells provide a valuable research tool to investigate cellular mechanisms in the pulmonary microvasculature relevant to the pathogenesis of emphysema. BioMed Central 2013 2013-02-20 /pmc/articles/PMC3599007/ /pubmed/23425195 http://dx.doi.org/10.1186/1465-9921-14-23 Text en Copyright ©2013 Mackay et al; licensee BioMed Central Ltd. http://creativecommons.org/licenses/by/2.0 This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. |
spellingShingle | Research Mackay, Laura S Dodd, Sara Dougall, Iain G Tomlinson, Wendy Lordan, James Fisher, Andrew J Corris, Paul A Isolation and characterisation of human pulmonary microvascular endothelial cells from patients with severe emphysema |
title | Isolation and characterisation of human pulmonary microvascular endothelial cells from patients with severe emphysema |
title_full | Isolation and characterisation of human pulmonary microvascular endothelial cells from patients with severe emphysema |
title_fullStr | Isolation and characterisation of human pulmonary microvascular endothelial cells from patients with severe emphysema |
title_full_unstemmed | Isolation and characterisation of human pulmonary microvascular endothelial cells from patients with severe emphysema |
title_short | Isolation and characterisation of human pulmonary microvascular endothelial cells from patients with severe emphysema |
title_sort | isolation and characterisation of human pulmonary microvascular endothelial cells from patients with severe emphysema |
topic | Research |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3599007/ https://www.ncbi.nlm.nih.gov/pubmed/23425195 http://dx.doi.org/10.1186/1465-9921-14-23 |
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